ALBERT

Description: Abnormal activation of kinases may promote leukemogenesis by conferring cell proliferation and survival advantage in acute myelogenous leukemia (AML). New targeted therapies are currently developed to disrupt such kinases activation to improve the long term prognosis of AML patients. However, the prognosis impact on AML patients has only been evaluated for a few kinases, including FLT3 and ERK/MAPK. Recent reports suggest that PI3K activation may confer poor survival to AML patients. Pre-treatment primary bone marrow blast cells samples from 92 patients registered for the LAM2001 trial of the French GOELAMS group were analyzed and PI3 kinase constitutive activation was correlated with outcome. As previously reported (V Bardet, Haematologica, 2006, 91, 757–764), the status of AKT phosphorylation on Ser 473 was determined using either Western Blot analysis (in samples 〉90% blasts) or flow cytometry in the CD45Low positive population of blast cells. Phosphorylation of AKT on Ser473 was determined after a 4 hour period of deprivation in cytokine and serum-free medium. In such conditions, we can clearly define to groups of patients, one with constitutive activation of PI3K (PI3K+) and the other without detectable phosphorylation of AKT (PI3K−). At inclusion, 42 patients were males and 50 were females. Median age at diagnosis was 46 (17,3–64,5) years. Leukocytosis was 12000/μL (range 400–251800). Patients were classified in low-risk, standard-risk and high-risk cytogenetic subgroups in respectively 16%, 61% and 23%. 16/61 (25%) had FLT3-ITD mutation, 9/60 (15%) had Ras (N-Ras or K-Ras) mutations and 15/59 (25%) had NPM mutations. 46 of 92 (50%) patients had constitutive PI3K activation (PI3K+ group). No difference was observed between the PI3K+ and PI3K− groups for all previously descripted parameters, except concerning FLT3 mutational status: 5/32 (16%) PI3K+ patients had FLT3-ITD versus 11/29 (38%) in the PI3K− (p = 0,048). All patients received one course of induction chemotherapy (3+7 regimen), and 31 (34%) a second induction course at day 15. Complete remission was obtained in 74 patients (80%) with no difference between the two groups (p = 0,60). Among responders, 30 (41%) had autologous stem cell transplantation, 24 (32%) had allogenic bone marrow transplantation and 16 (22%) had consolidation chemotherapy. With a median follow-up of 26 months (range 3 days–37 months), 48 patients died. Overall survival was 56% in the PI3K+ group and 33% in the PI3K− group (p=0.007). Among responders, 30 patients had relapse. Relapse free survival was 72% in the PI3K+ group and 41% in the PI3K− group (p=0.001). Thus constitutive PI3K activity detected in leukemic cells at diagnosis confer a better prognosis in AML. We observed significantly more relapses in PI3K− patients. These results suggest that PI3K activity in AML may confer a particular sensitivity to chemotherapy, the demonstration of which remain to be made. Furthermore, inhibition of PI3K signaling pathway may not be as beneficial as expected, and a better understanding of downstream effectors is clearly needed before targeting PI3K in AML.

Description: Introduction. The prognosis of AML in patients over 60 years of age remains very poor. As in younger patients karyotype is a major prognostic factor. Elderly patients with poor risk cytogenetics have a dismal outcome and complete remission (CR) rates with conventional chemotherapy are usually 〈 45% (and 〈 25% if complex abnormalities). GO has been used in combination with chemotherapy in relapsed or newly diagnosed AML, but with reduced doses in order to decrease hepatic toxicity. Methods. In this multicentric open-label Phase II study we have evaluated the combination of I (8 mg/m2/D × D1-5) + C (100 mg/m2/D × D1-7) with GO (6 mg/m2 on D3) in AML patients aged 60–75 years with poor risk cytogenetics and fit enough to undergo cytotoxic chemotherapy. Other inclusion criteria were: CD33+ AML, PS 2, de novo or secondary AML (excluding M3 and Ph1 + AML), no cardiac, renal or hepatic dysfunction. Results. 44 pts aged 60–74 (median 68) were treated in 13 GOELAMS centers. Karyotype results were: chromosome (chr) 5 abnormalities 22, chr 7 abnormalities 26, chr 5+7 abnormalities 14, complex abnormalities 26. In 13 cases (30%) AML was secondary (chemoinduced 9, prior myeloproliferative or myelodysplastic syndrome 4). Median duration of hospitalisation was 31 D (4–51). In CR patients median duration of neutropenia was 19 D (14–28) and median duration of thrombocytopenia was 15D (5–29). Bacterial or fungal infections were documented in 71% of cases. 7 patients had severe bleeding including 3 deaths, (2 cerebral hemorrhage, 1 intestinal bleeding). Grade 2 hepatic dysfunction was seen in 30% of patients including 3 (7%) pts with veino-occlusive disease (one death). There were 17 (38%) CR (16 CR, 1 CRp), 6 (14%) toxic deaths and 21 (48%) failures. The CR rates was 41% (7/17) for patients aged 70–75 and 37% (10/27) for patients 〈 70 years. While CR rate was 50% (6/12) in patients with isolated chr 5 or 7 abnormalities, it was 35% (9/26) in patients with complex abnormalities. Conclusion. In this population of elderly but relatively fit patients with poor risk cytogenetics AML the addition of GO 6 mg/m2 to conventional chemotherapy did not appear to increase the CR rate mostly because of leukemic resistance. The incidence of aplasia-related toxicities and of hepatic complications does not encourage to use higher doses of GO.

Description: Background: In recent years, oncogenic understanding of T-ALL has led to the identification of multiple molecular markers. However, each molecular abnormality accounts for a small proportion of cases and risk stratification at diagnosis still relies on age, clinical presentation, and early response to therapy. NOTCH1 and/or FBXW7 mutations have been recently reported to be a recurrent abnormality in T-ALL, both leading to activation of the NOTCH pathway. Studies in pediatric series have suggested a favorable outcome for NOTCH1 mutated T-ALL, but this has not been evaluated in large series of adult cases. Furthermore, FBXW7 prognostic impact remains unknown in both populations. Methods: In order to evaluate the incidence of these mutations and their prognostic impact in adults, we performed a retrospective analysis of 141 patients (median age, 28 years) with T-ALL treated within the LALA-94 (N=87) or the more recent GRAALL-2003 (N=54) French trials. These patients were representative of the overall population since their outcome did not differ from the overall T-ALL cases treated in either LALA-94 (estimated 3-year OS, 41%) or GRAALL-2003 (estimated 3-year OS, 66%) trial. Furthermore, the patients from the LALA-94 and the GRAALL-2003 trials did not differ with respect to sex ratio, age, WBC, and initial complete remission rate (92% and 98%, respectively). Exons 26 (HD N-terminal), 27 (HD C-terminal), 28 (juxtamembrane domain) and 34 (transactivation domain TAD and the PEST domain) of NOTCH1 and exons 9, 10 and 12 of WD40 domain of FBXW7 were sequenced. Results: We identified 101 cases with NOTCH1 and/or FBXW7 mutations (72%) and 40 wild type (WT) samples (28%). NOTCH1 was mutated in 88 patients (59 HD only, 15 HD+PEST, 9 PEST only, 5 other). FBXW7 was mutated in 34 patients, alone in 13 cases or in association with NOTCH1 mutations in 21 cases. There was no significant correlation between NOTCH1 and/or FBXW7 mutations and immunophenotypic or oncogenetic features. There was a trend for a higher WBC and more frequent CNS involvement in patients demonstrating WT NOTCH1 and FBXW7. Similarly, high-risk MRC/ECOG criteria (age〉35y and/or WBC〉100G/L) were found in 56% of patients from the mutated subgroup versus 73% in the WT subgroup (P=0.06). The prognostic impact of NOTCH1 mutations alone did not reach statistical significance on multivariate analysis (P=0.09). On the other hand, multivariate analysis showed that the GRAALL-2003 trial and the presence of NOTCH1 and/or FBXW7 mutations were the only factors associated with a longer EFS (P=0.001 and 0.035, respectively). Median EFS was 36 months for patients with NOTCH1 and/or FBXW7 mutations versus 17 months for WT patients. Conclusions: These data demonstrate that NOTCH pathway activation by either NOTCH1 or FBXW7 mutation identifies a large group of T-ALL adult patients (72%) with a favorable outcome that could be used for treatment stratification.

Description: Ph+ ALL accounts for approximately one third of ALL cases in patients aged 55 years or older. The median survival of older Ph+ ALL patients is one year, with practically no long-term survivor (Blood, 98, Supp1 p319a, 2001). Imatinib has demonstrated remarkable, although transient, activity in relapsed and refractory Ph+ ALL, which prompted the GRAALL to implement a treatment protocol associating imatinib and chemotherapy in previously untreated elderly patients: ALL patients aged 55 years or older are treated with steroids during one week and Ph+ve cases are then offered a specific therapy including an induction treatment with steroids, cyclophosphamide, daunorubicin and vincristine, followed, irrespective of response to induction chemotherapy, by imatinib, 600 mg daily, combined with intermittent steroids during 2 months. Patients in complete response are then given 10 blocks of alternating chemotherapy, including 2 additional two-month blocks of imatinib, for a total treatment duration of 2 years. Therapy of occult central nervous system leukemia includes 5 intrathecal injections of methotrexate and cranial irradiation. The study is intended to include 30 patients and its main objective is to improve overall one-year survival to 70%. Results are compared with those obtained in 21 Ph+ ALL elderly patients treated according to our previous protocol. Since January 2003, 21 patients aged 58 to 78 years (median: 64.7 years) were included in the AFR09 protocol. Their median follow-up is 3 months. 15/19 patients are in complete response after induction chemotherapy vs 6/21 in the historical controls given similar induction regimen but with no steroids before chemotherapy (p=0.002). The projected overall survival is 95% at 9 months vs 62% in the control group (p=0.08, log-rank test). The 9-month projected event-free survival is 83% vs 10% (p

Description: In a large series of patients with Ph1-ALL treated in the LALA-94 trial, negative BCR-ABL minimal residual disease (MRD) and allogeneic donor availability were two independent good-risk factors for survival in those reaching hematological complete remission (HCR) after standard induction followed by intensive HAM consolidation (Dombret et al. Blood2002;100:2357). The HAM regimen included intermediate-dose cytarabine (1 g/m2/12h Day 1 to 4) and mitoxantrone (10 mg/m2 Day 3 to 5). The fraction of HCR patients in molecular CR (MCR) after HAM was 38%, with a median RT-PCR sensitivity of 10−5. In the present Phase I/II study, we combined increasing dosages of imatinib with HAM (HAMI regimen) in an attempt: 1) to increase the fraction of MCR patients from 38 to 75% (as primary objective); and 2) to evaluate the safety of the combination regimen and the feasibility of autologous peripheral blood stem cell (PBSC) collection after HAMI (as secondary objectives). Patients aged 18–60 years with Ph1-ALL in HCR after 1 or 2 cycles of chemotherapy were eligible in the absence of counter-indication for further stem cell transplantation. Imatinib was administered daily from Day 1 of HAM. MRD was assessed by RQ-PCR at Day 0 and Day 45 of HAMI. MCR was defined as a BCR-ABL level lower than 10−5 at Day 45. Twenty-two patients have been enrolled. Results observed in the first 18 patients treated in 3 consecutive 6-patient cohorts at 400, 600, and 800 mg/d imatinib, respectively, are available. Main results may be summarized as follows: 1) an excessive toxicity was observed in the 800 mg/d cohort leading to recommend the 600 mg/d dosage for further combinations of imatinib and intensive chemotherapy; 2) only 5 of the 15 patients tested (33%) reached MCR at Day 45 of HAMI and no clear correlations were found between molecular response and imatinib dosage or Day 0 MRD level; 3) after HAMI, PBSC collection was feasible under imatinib administration in 9 out of 14 patients (64%) with an apparent relationship between failure to collect and no MCR (P=.08); 4) only 3 patients were not able to receive allogeneic or autologous transplantation as further consolidation (1 early relapse, 1 severe infection, 1 too high MRD level); and 5) estimated 18-month disease-free and overall survival were 58% and 78%, respectively, with 8/15 patients receiving imatinib during the post-transplant period. In conclusion, failure to improve the MCR rate as compared to our no-imatinib historical control could be explained by the relatively short exposure to imatinib and/or by the absence of in vivo synergism between imatinib and cytarabine or mitoxantrone. More benefit might be anticipated by incorporating imatinib earlier in the treament i.e. first induction course. Further studies are needed to evaluate the role of post-transplant maintenance with imatinib.

Description: The incidence of AT and Fg deficiency and of thrombotic and bleeding events after Asp has been well evaluated in children with ALL, but little is known in adults. In this study, the incidence of these events and the use of coagulation supportive treatments was evaluated in 214 adult patients with ALL (n= 191) or LBL (n=23) included in the GOELAL02 trial (Blood 2004, in press). The induction therapy included steroids (40 mg/m2/d d1-21), vincristine (2 mg d1, 8, 15, 22), idarubicine (5 mg/m2 d1, 8,15, 22) and Asp (7500 UI/m2 d10, 13, 16, 19, 22, 25) delivered through a central venous access. Fresh frozen plasma (FFP), Fg (Clottagen®) and AT (Aclotine®) concentrates were recommended to maintain their levels 〉 1 g/l and 60%, respectively. If no transfusion, Asp infusion was delayed for 48 hours. Platelets were transfused when 〈 20 x 109/l. Heparin prophylaxis was left to institutional guidelines. Symptomatic thromboses and significant bleedings were systematically recorded. Active DIC was present in 10.3% of patients on d1. Fg and AT levels were measured 4098 times (20/patient, von Clauss assay) and 1718 times (8/patient, chromogenic assay) respectively, and evolved between d1 and d35 as shown below (mean ± sd). AT nadir was 〈 60% in 71% of patients with values 〈 40% in 26% of cases. Fg levels were 〈 1g/l in 73% of patients with values 〈 0.5g/l in 9%.Twenty-one thromboses occurred two to 35d after the first Asp infusion (median = 14d) in 20 patients (9.3%), with cerebral vein thrombosis (5), pulmonary embolism (3), upper (5) or lower (8) limb deep vein thrombosis. At the time of event, the median AT level was 48% (mean 60.7) with 12 of 21 thromboses (57%) occurring with AT 〈 60%. Fourty-two bleedings occurred one to 45d after the first Asp infusion (median = 8d) in 31 patients, with CNS hemorrhage (1), epistaxis (24), central venous access bleeding (8), GI bleeding (1), and large ecchymoses (8). At the time of event, median Fg level was 1.3 g/l. Asp infusions were reduced or delayed in 64% of all patients due to low Fg and/or AT levels. FFP, AT and Fg were infused in 31%, 41% and 52% of patients at mean doses of 5.3 ml/kg/infusion, 31UI/kg/infusion and 7.9 g respectively. AT level increased from 58%±16 (n=79) to 86%±23.2 (n=57) after the first AT infusion but was unchanged after FFP. Fg level increased from 0.9g/l ± 0.3 (n=85) to 1.4g/l ±0.5 (n=69) after Fg infusion but was unmodified after FFP. In conclusion, Fg and AT levels are frequently decreased in adults treated by Asp, with often a less than optimal chemotherapy. Bleeding events were not associated with severe Fg deficiency, but thrombotic events could be favored in some patients by acquired AT deficiency. The benefit of AT concentrates to prevent thrombosis and to reduce delay in Asp infusions could therefore be prospectively assessed in adults treated by Asp. d1 (n) d10-Asp 1 (n) d13-Asp2 (n) d19-Asp 4 (n) d25-Asp 6 (n) AT % - 121±16 (79) 83±16 (125) 65±20 (111) 65±22 (77) Fg g/l 3.2±1.7 (142) 1.9±1.0 (159) 1.4±0.7 (195) 1.1±0.4 (169) 1.4±0.6 (118)

Description: Many somatic genetic abnormalities have been identified in T-cell acute lymphoblastic leukemia (T-ALL) but each individual abnormality accounts for a small proportion of cases; therapeutic stratification consequently still relies on classical clinical markers. NOTCH1 and/or FBXW7 mutations both lead to activation of the NOTCH1 pathway and are among the most frequent mutations in T-ALL. We screened 141 adult diagnostic T-ALL samples from patients treated on either the Lymphoblastic Acute Leukemia in Adults (LALA)-94 (n = 87) or the GRAALL-2003 (n = 54) trials. In 88 cases (62%) there were demonstrated NOTCH1 mutations (42% heterodimerization [HD], 10% HD+proline glutamate serine threonine [PEST], 6% PEST, 2% juxtamembrane mutations, 2% transactivation domain [TAD]) and 34 cases (24%) had FBXW7 mutations (21 cases had both NOTCH1 and FBXW7 mutations); 40 cases (28%) were wild type for both. There was no significant correlation between NOTCH1 and/or FBXW7 mutations and clinico-biologic features. Median event-free survival (EFS) and overall survival (OS) were 36 versus 17 months (P = .01) and not reached versus 32 months (P = .004) in patients with NOTCH1 and/or FBXW7 mutations versus other patients, respectively. Multivariate analysis showed that the presence of NOTCH1/FBXW7 mutations was an independent good prognostic factor for EFS and OS (P = .02 and P = .01, respectively). These data demonstrate that NOTCH1 pathway activation by either NOTCH1 or FBXW7 mutation identifies a large group of patients with a favorable outcome that could justify individual therapeutic stratification for T-ALL.

Description: The prognosis of acute myeloid leukemia (AML) is very poor in elderly patients, especially in those classically defined as having unfavorable cytogenetics. The recent monosomal karyotype (MK) entity, defined as 2 or more autosomal monosomies or combination of 1 monosomy with structural abnormalities, has been reported to be associated with a worse outcome than the traditional complex karyotype (CK). In this retrospective study of 186 AML patients older than 60 years, the prognostic influence of MK was used to further stratify elderly patients with unfavorable cytogenetics. CK was observed in 129 patients (69%), and 110 exhibited abnormalities according to the definition of MK (59%). MK+ patients had a complete response rate significantly lower than MK− patients: 37% vs 64% (P = .0008), and their 2-year overall survival was also decreased at 7% vs 22% (P 〈 .0001). In multivariate analysis, MK appeared as the major independent prognostic factor related to complete remission achievement (odds ratio = 2.3; 95% confidence interval, 1-5.4, P = .05) and survival (hazard ratio = 1.7; 95% confidence interval, 1.1-2.5, P = .008). In the subgroup of 129 CK+ patients, survival was dramatically decreased for MK+ patients (8% vs 28% at P = .03). These results demonstrate that MK is a major independent factor of very poor prognosis in elderly AML.

Description: In spite of the tremendous progresses of therapeutic approaches in the past decades, the prognosis of AML remains very poor. Developing new biological tools for disease monitoring is therefore a constant challenge, to adapt therapeutic protocols as early as possible. It has been suggested (Leuk Res,1992) that a morphologically assessed rate of decrease of medullary blasts during induction could be a prognostic indicator. Increasing sophistication of flow cytometry (FC) has considerably improved the sensitivity of this technique, allowing for a precise and reliable detection of low blast percentages (

Description: Karyotype is one of the most important prognosis factor in acute myeloid leukemia. The MLL gene, located at 11q23, is fused to a variety of partner genes through chromosomal translocations and generates the critical leukemogenic fusion proteins. We performed a large retrospective study in 146 adults and children with 11q23/MLL + rearrangement enrolled in 6 clinical trials conducted in France between 1987 and 1998. Prognosis studies were performed in the 110 patients (47 adults and 63 children) who achieved a complete remission. Individual data were registred, including age, blood and marrow count, extramedullary disease and cytogenetics. The kaplan-Meier method was used in survival analysis and the cox proportinal-hazards model was used to analyse the effect of potential prognosis factors on survival. Among 47 adults, 43 % (20/47) carried t(9 ;11), 19% (9/47) carried t(6 ;11), 23% (11/47) carried t(11 ;19), 6% (3/47) carried t(10 ;11) and 8% (4/47) carried another abnormalities involved 11q23/MLL. Estimated 5-year disease-free survival (DFS), overall survival (OS) and relapse rate (RR) were respectively 17%, 21% and 77%. We analysed separately all translocations involved MLL and we found that the only translocation indicating a favorable prognosis was the translocation t(11 ;19). Compared with other translocations involving MLL, the estimated 5-year DFS, OS and RR of patients with a t(11 ;19) were respectively 45 vs 8% (p=.02), 54 vs 11% (p=.006) and 48 vs 84% (p=.04). Relative risk of relapse was 0,29 (95% confidence interval, 0.09 to 0,9 ;p=.03). Among 63 children, 63% (40/63) carried t(9 ;11), 3% (2/63) carried t(6 ;11), 8% (5/63) carried t(11 ;19), 14% (9/63) carried t(10 ;11) and 11% (7/63) carried another MLL rearrangement. Estimated 5-year DFS, OS and RR were respectively 59%, 72% and 36%. Compared with other translocations involving MLL, we found no difference in outcome between patients with t(9 ;11) and those with other rearrangement of MLL. In conclusion, the most common recurrent abnormality in this large study was translocation t(9 ;11) in adults and children. We found that adults patients whose leukemic cells contained the t(11 ;19) have a better outcome compared with those whose leukemic cells contain other 11q23 /MLL alterations. In children, we do not confirm the favorable impact of t(9 ;11) suggested in previous reports.